This chapter provides an overview of class B β-lactamases (CBBLs), considering both fundamental and clinical aspects. Class B is one of the four classes in the structural classification of β-lactamases, which was created to accommodate the metallo-β-lactamases (MBLs). Expression of the CBBL genes carried on gene cassettes is normally under the control of the integron promoters (Pc and, possibly, P2) located in the 5'-conserved segment of the integron. The constant features of CBBLs include (i) good to excellent carbapenemase activity; (ii) lack of activity on monobactams, which apparently do not interact with these enzymes; and (iii) inhibition by EDTA and other metal ion chelators, and lack of inhibition by the conventional serine-β-lactamase inhibitors. Concerning the structure of the zinc center, which is located at the bottom of a shallow groove between the two β-sheets, in subclass B1 and B3 CBBLs it can accommodate two zinc ions (dinuclear zinc center): a tetrahedrally coordinated zinc ion (Zn1) and a trigonal bipyramidally coordinated zinc ion (Zn2), bridged by a water molecule/hydroxide ion (Wat1). Zn1 is coordinated by three His residues (His 116, His118, and His196) and the bridging Wat1 in all subclass B1 and B3 enzymes, while the strategy of Zn2 coordination is partially different in members of the two subclasses. The clinical relevance of resident CBBLs essentially reflects that of the host species, where these enzymes can variably contribute to intrinsic β-lactam resistance depending on their expression pattern and substrate specificity.

Unrooted tree showing the structural relatedness among the major lineages of CBBLs. The names of the enzymes are the same as in Table 9.1. The tree was constructed using the TREEVIEW program (121) on the basis of a sequence alignment constructed with the CLUSTAL X program (158) using the sequences whose accession numbers are reported in the fourth column of Table 9.1.

10.1128/9781555815615/f0118-01_thmb.gif

10.1128/9781555815615/f0118-01.gif

Figure 9.1

Unrooted tree showing the structural relatedness among the major lineages of CBBLs. The names of the enzymes are the same as in Table 9.1. The tree was constructed using the TREEVIEW program (121) on the basis of a sequence alignment constructed with the CLUSTAL X program (158) using the sequences whose accession numbers are reported in the fourth column of Table 9.1.

Amino acid sequence alignment of CBBLs of subclass B1 (section A), subclass B2 (section B), and subclass B3 (section C). The alignments were constructed considering a single representative for each major lineage, using the sequences whose accession numbers are reported in the fourth column of Table 9.1. Structural elements are shown above the alignment for enzymes of each subclass, based on the three-dimensional structures of the Bc-II (23), CphA (53), and L1 (167) enzymes, respectively. The conserved residues shared by enzymes of subclass B1 and those shared by enzymes of subclass B3 are boxshaded in gray (note that the Trp244 is not conserved in IMP-18). The four residues conserved among all CBBLs are boxshaded in black. Numbering is according to the BBL scheme (52, 54).

10.1128/9781555815615/f0119-01_thmb.gif

10.1128/9781555815615/f0119-01.gif

Figure 9.2

Amino acid sequence alignment of CBBLs of subclass B1 (section A), subclass B2 (section B), and subclass B3 (section C). The alignments were constructed considering a single representative for each major lineage, using the sequences whose accession numbers are reported in the fourth column of Table 9.1. Structural elements are shown above the alignment for enzymes of each subclass, based on the three-dimensional structures of the Bc-II (23), CphA (53), and L1 (167) enzymes, respectively. The conserved residues shared by enzymes of subclass B1 and those shared by enzymes of subclass B3 are boxshaded in gray (note that the Trp244 is not conserved in IMP-18). The four residues conserved among all CBBLs are boxshaded in black. Numbering is according to the BBL scheme (52, 54).

Unrooted tree showing the structural relatedness among known IMP variants. The tree was constructed using the TREEVIEW program (121) on the basis of a sequence alignment constructed with the CLUSTAL X program (158) using the sequences whose accession numbers are reported in the third column of Table 9.2.

10.1128/9781555815615/f0121-01_thmb.gif

10.1128/9781555815615/f0121-01.gif

Figure 9.3

Unrooted tree showing the structural relatedness among known IMP variants. The tree was constructed using the TREEVIEW program (121) on the basis of a sequence alignment constructed with the CLUSTAL X program (158) using the sequences whose accession numbers are reported in the third column of Table 9.2.

Unrooted tree showing the structural relatedness among known VIM variants. The tree was constructed using the TREEVIEW program (121) on the basis of a sequence alignment constructed with the CLUSTAL X program (158) using the sequences whose accession numbers are reported in the third column of Table 9.3.

10.1128/9781555815615/f0123-01_thmb.gif

10.1128/9781555815615/f0123-01.gif

Figure 9.4

Unrooted tree showing the structural relatedness among known VIM variants. The tree was constructed using the TREEVIEW program (121) on the basis of a sequence alignment constructed with the CLUSTAL X program (158) using the sequences whose accession numbers are reported in the third column of Table 9.3.

Schematic representation of the structure of the variable region of class 1 integrons containing blaIMP (section A) or blaVIM(section B) gene cassettes. The gene cassettes are indicated by arrows (those carrying CBBL genes are filled). The presence of the integron 5′-conserved segment (5′-CS) and 3′-conserved segment (3′-CS), flanking the cassette array, is also indicated. Only integrons whose variable region has been completely sequenced are shown.

10.1128/9781555815615/f0124-01_thmb.gif

10.1128/9781555815615/f0124-01.gif

Figure 9.5a

Schematic representation of the structure of the variable region of class 1 integrons containing blaIMP (section A) or blaVIM(section B) gene cassettes. The gene cassettes are indicated by arrows (those carrying CBBL genes are filled). The presence of the integron 5′-conserved segment (5′-CS) and 3′-conserved segment (3′-CS), flanking the cassette array, is also indicated. Only integrons whose variable region has been completely sequenced are shown.

Schematic representation of the structure of the variable region of class 1 integrons containing blaIMP (section A) or blaVIM(section B) gene cassettes. The gene cassettes are indicated by arrows (those carrying CBBL genes are filled). The presence of the integron 5′-conserved segment (5′-CS) and 3′-conserved segment (3′-CS), flanking the cassette array, is also indicated. Only integrons whose variable region has been completely sequenced are shown.

10.1128/9781555815615/f0125-01_thmb.gif

10.1128/9781555815615/f0125-01.gif

Figure 9.5b

Schematic representation of the structure of the variable region of class 1 integrons containing blaIMP (section A) or blaVIM(section B) gene cassettes. The gene cassettes are indicated by arrows (those carrying CBBL genes are filled). The presence of the integron 5′-conserved segment (5′-CS) and 3′-conserved segment (3′-CS), flanking the cassette array, is also indicated. Only integrons whose variable region has been completely sequenced are shown.

Schematic representation of the structure of the variable region of class 1 integrons containing blaIMP (section A) or blaVIM(section B) gene cassettes. The gene cassettes are indicated by arrows (those carrying CBBL genes are filled). The presence of the integron 5′-conserved segment (5′-CS) and 3′-conserved segment (3′-CS), flanking the cassette array, is also indicated. Only integrons whose variable region has been completely sequenced are shown.

10.1128/9781555815615/f0126-01_thmb.gif

10.1128/9781555815615/f0126-01.gif

Figure 9.5c

Schematic representation of the structure of the variable region of class 1 integrons containing blaIMP (section A) or blaVIM(section B) gene cassettes. The gene cassettes are indicated by arrows (those carrying CBBL genes are filled). The presence of the integron 5′-conserved segment (5′-CS) and 3′-conserved segment (3′-CS), flanking the cassette array, is also indicated. Only integrons whose variable region has been completely sequenced are shown.

(Upper panels) Ribbon diagram of the three-dimensional structure of the CfiA/CcrA enzyme from B. fragilis strain QMCN3 (PDB accession no., 1ZNB), of the CphA enzyme from A. hydrophila AE036 (PDB accession no., 1X8G), and of the L1 enzyme from S. maltophilia strain IID 1275 (PDB accession, 1SML). The diagrams were constructed using the MOLMOL program (86). (Lower panels) Structure of the zinc centers of the corresponding enzymes (the water molecule is replaced by a carbonate in the CphA structure, and a single oxygen atom, involved in zinc coordination, is shown for clarity).

10.1128/9781555815615/f0131-01_thmb.gif

10.1128/9781555815615/f0131-01.gif

Figure 9.6

(Upper panels) Ribbon diagram of the three-dimensional structure of the CfiA/CcrA enzyme from B. fragilis strain QMCN3 (PDB accession no., 1ZNB), of the CphA enzyme from A. hydrophila AE036 (PDB accession no., 1X8G), and of the L1 enzyme from S. maltophilia strain IID 1275 (PDB accession, 1SML). The diagrams were constructed using the MOLMOL program (86). (Lower panels) Structure of the zinc centers of the corresponding enzymes (the water molecule is replaced by a carbonate in the CphA structure, and a single oxygen atom, involved in zinc coordination, is shown for clarity).

130. Podglajen,I.,, J.Breuil,, I.Casin, and, E.Collatz.1995.Genotypic identification of two groups within the species Bacteroides fragilis by ribotyping and by analysis of PCR-generated fragment patterns and insertion sequence content.J. Bacteriol.177:5270–5275.

148. Rossolini,G. M.,, A.Zanchi,, A.Chiesurin,, G.Amicosante,, G.Satta, and, P.Guglielmetti.1995.Distribution of cphA or related carbapenemase-encoding genes and production of carbapenemase activity in members of the genus Aero-monas.Antimicrob. Agents Chemother.39:346–349.